Host defense peptide LEAP-2 contributes to monocyte/macrophage polarization in barbel steed (Hemibarbus labeo).

The liver-expressed antimicrobial peptide 2 (LEAP-2) plays a vital role in host immunity against pathogenic organisms. In the present study, cDNA of the LEAP-2 gene was cloned and sequenced from the barbel steed (Hemibarbus labeo). The predicted amino acid sequence of the barbel steed LEAP-2 comprises a signal peptide and a prodomain, which is followed by the mature peptide. Sequence analysis revealed that barbel steed LEAP-2 belongs to the fish LEAP-2A cluster and that it is closely related to zebrafish LEAP-2A. We found that barbel steed LEAP-2 transcripts were expressed in a wide range of tissues, with the highest mRNA levels detected in the liver. In response to lipopolysaccharide (LPS) treatment, LEAP-2 was significantly upregulated in the liver, head kidney, spleen, gill, and mid intestine. A chemically synthesized LEAP-2 mature peptide exhibited selective antimicrobial activity against several bacteria in vitro. Moreover, LEAP-2, alone or in combination with LPS or phorbol 12-myristate 13-acetate, strongly induced a pro-inflammatory reaction in barbel steed monocytes/macrophages (MO/MФ), involving the induction of iNOS activity, respiratory burst, and the pro-inflammatory cytokines IFN-γ, TNF-α, and IL-1ß. Collectively, the results of this study indicate the importance of fish LEAP-2 in the M1-type polarization of MO/MΦ.

Molecular characterization of a hepcidin homologue in starry flounder (Platichthys stellatus) and its synergistic interaction with antibiotics.

Hepcidins are small cysteine-rich antimicrobial peptides that play an important role in host immunity against pathogenic organisms. Most fish hepcidins exert bactericidal activities against a wide range of pathogens. In this study, we identified a cDNA sequence encoding a hepcidin homologue (PsHepcidin) in the starry flounder Platichthys stellatus. The predicted amino acid sequence of PsHepcidin comprises a signal peptide and a prodomain, which are followed by the mature peptide. Sequence analysis revealed that PsHepcidin belongs to the fish HAMP2 cluster and that it is closely related to mudskipper hepcidin-2. Expression of PsHepcidin mRNA was detected in all examined immune-related tissues, with the highest transcript levels being found in the liver. In response to lipopolysaccharide treatment, PsHepcidin was significantly up-regulated in the liver, kidney, and spleen in a time-dependent manner. Chemically synthesized mature peptides of PsHepcidin were found to exhibit broad antimicrobial activity in vitro. We also investigated the combined effect of PsHepcidin and conventional antibiotics and found that these combinations showed synergistic effects against most of the examined bacterial strains. Collectively, the results of this study indicate that PsHepcidin exhibits potent antibacterial activity both independently and when used in combination with conventional antibiotics.

Effect of high temperature stress on heat shock protein expression and antioxidant enzyme activity of two morphs of the mud crab Scylla paramamosain.

The present study aimed to investigate the effect rapid temperature change from moderate temperature to high temperatures on heat shock protein (HSP) expression and antioxidant enzyme activities in mud crabs. Two mud crabs, one with one spine on the outer margin of the carpus of cheliped (Sp1) and another with two spines (Sp2), were acclimated at 25 °C and then transferred to a 33 °C environment, and HSP expression and antioxidant enzyme activity were assessed. HSP70 and HSP60 were markedly up-regulated in the gills and hepatopancreas of Sp1 and Sp2 after exposure to 35 °C. Exposure to 35 °C also significantly increased superoxide dismutase and catalase activity in the gills of Sp1 and Sp2, with transient changes in hepatopancreas. Apart from changes in antioxidant enzyme activities, HSPs were highly up-regulated after exposure to 37 °C, especially for HSP70. Gill HSP70 expression in Sp2 was 6.1 folds that of the control after 24 h of exposure to 37 °C, and 9.2 folds that of Sp1. Moreover, exposure to 37 °C further up-regulated HSP70 in the hepatopancreas of Sp1, compared to that in Sp2. Hence, HSPs play important roles in thermotolerance in S. paramamosain and Sp1 might have a stronger tolerance to hyperthermal stress than Sp2.